首先在此感谢jimmy非常详尽的教程 HiC数据分析实战之HiC-Pro
本文为三维基因组学习笔记的第二篇，主要记录HiC-pro的安装遇到的问题及部分实战。
安装
- 首先根据要求说明安装依赖，可由conda安装，注意版本
- The bowtie2 mapper
- Python (>2.7, python-3 is not supported) with pysam (>=0.8.3), bx-python(>=0.5.0), numpy(>=1.8.2), and scipy(>=0.15.1)libraries
- R with the RColorBrewer and ggplot2 (>2.2.1) packages
- g++ compiler
- samtools (>1.1)
- Unix sort (which support -V option) is required ! For Mac OS user, please install the GNU core utilities !
- HiC-Pro的安装
不在conda中的软件代码
$ pip install https://bitbucket.org/mirnylab/mirnylib/get/tip.tar.gz
$ pip install https://bitbucket.org/mirnylab/hiclib/get/tip.tar.gz
# hicpro的安装
$ cd ~/biosoft/hicpro
$ cd ~/biosoft/hicpro
$ git clone https://github.com/nservant/HiC-Pro.git
$ cd HiC-Pro
# 这里要改写配置文件（见下）
$ cat config-install.txt
$ mkdir ~/biosoft/hicpro/bin
$ make configure
$ make install
### 最后安装的时候可能会出现Directory does not exit!,这可能是程序默认在home目录下有bin这个文件夹导致，新建bin文件夹即可。最后 绝对路径/HiC-Pro -h跳出说明即安装成功。
SYSTEM CONFIGURATION
| PREFIX | Path to installation folder |
|---|---|
| BOWTIE2_PATH | Full path the bowtie2 installation directory |
| SAMTOOLS_PATH | Full path to the samtools installation directory |
| R_PATH | Full path to the R installation directory |
| PYTHON_PATH | Full path to the python installation directory (>2.7 - python3 not supported) |
| CLUSTER_SYS | Scheduler to use for cluster submission. Must be TORQUE, SGE, SLURM or LSF |
运行
- 首先需要获得消化片段的BED文件及chromosomes' size表格文件，这里需要限制酶酶切位点及参考基因组信息。根据测试数据来源及digest_genome.py
$ /PATH/HiC-Pro-master/bin/utils/digest_genome.py -r hindiii -o Refgenome.fasta
# BED文件格式（-1）
chr1 0 16007 HIC_chr1_1 0 +
# chromosomes' size（-1）
chr1 249250621
HiC-Pro --help
usage : HiC-Pro -i INPUT -o OUTPUT -c CONFIG [-s ANALYSIS_STEP] [-p] [-h] [-v]
Use option -h|--help for more information
HiC-Pro 2.10.0
---------------
OPTIONS
-i|--input INPUT : input data folder; Must contains a folder per sample with input files
-o|--output OUTPUT : output folder
-c|--conf CONFIG : configuration file for Hi-C processing
[-p|--parallel] : if specified run HiC-Pro on a cluster
[-s|--step ANALYSIS_STEP] : run only a subset of the HiC-Pro workflow; if not specified the complete workflow is run
mapping: perform reads alignment
proc_hic: perform Hi-C filtering
quality_checks: run Hi-C quality control plots
build_contact_maps: build raw inter/intrachromosomal contact maps
ice_norm: run ICE normalization on contact maps
[-h|--help]: help
[-v|--version]: version
- 根据说明文档，将 configuration file 'config-hicpro.txt' 复制到你的当前目录，并修改；本次测试数据来源于来自于Tung B. K. Le et al. Science 2013 ：https://www.ncbi.nlm.nih.gov/sra/?term=srr824846，rawdata文件并不用编排，但是由于程序读写要求，因此需要将数据放入独立的文件夹中。
Put all input files in a rawdata folder. The input files have to be organized with one folder per sample, with ;
$ mkdir -p ~/data/project/hic/fq/s1/
$ cd ~/data/project/hic/fq/s1/
858M Jul 3 16:21 SRR824846_Q20L10_1.fastq.gz
857M Jul 3 16:22 SRR824846_Q20L10_2.fastq.gz
# 多个输入文件
+ PATH_TO_MY_DATA
+ sample1
++ file1_R1.fastq.gz
++ file1_R2.fastq.gz
++ ...
+ sample2
++ file1_R1.fastq.gz
++ file1_R2.fastq.gz
*...
- 运行命令如下，jimmy在推文中运用了一系列技巧，可以详细查看学习。
# 配置文件主要修改内容
BOWTIE2_IDX_PATH = # bowtie2建立的索引所在的路径，记住绝对路径
REFERENCE_GENOME = # bowtie2建立的索引
GENOME_SIZE = # 一个文件记录着参考基因组中每条序列的大小
GENOME_FRAGMENT = 消化片段的BED文件所在的路径
LIGATION_SITE = #连接位点
# 若单个测序数据则
PAIR1_EXT = SRR824846_Q20L10_1
PAIR2_EXT = SRR824846_Q20L10_2
$ MY_INSTALL_PATH/bin/HiC-Pro -i FULL_PATH_TO_DATA_FOLDER -o FULL_PATH_TO_OUTPUTS -c MY_LOCAL_CONFIG_FILE
$ cd out
$ qsub HiCPro_step1_.sh
$ qsub HiCPro_step2_.sh
这里记录一个问题，qsub命令报错，目前使用sh命令执行shell脚本。
$ qsub HiCPro_step1_.sh -p 20
Unable to initialize environment because of error: cell directory "/opt/gridengine/default" doesn't exist
Exiting.
待结果出来以后，将进一步学习。
相关链接地址：
HiC-Pro_github
HiC-Pro: An optimized and flexible pipeline for Hi-C processing
